Cyclic peptide automated synthesis

Robey, F. A. and Fields, R. L. (1989) Automated synthesis of bromoacetyl-modi-fied peptides for preparation of synthetic peptide polymers, peptide-protein conjugates and cyclic peptides. Anal. Biochem. 177, 373-377. 

Recently, cLac (cyclic peptide-mimicking lactadherin) was synthesized using NCL and studied for phosphatidylserine (PS) recognition [64, 65]. The linear precursors for the synthesis of cLac derivatives were prepared following standard protocols for automated Fmoc-peptide synthesis and the cyclic peptides (cLac variants) were obtained in the presence of 4-mercaptophenylacetic acid and isolated via HPLC in yields of 60-70% (Fig. 9). All the synthesized cLac variants were labeled with the thiol-reactive fluorescein-5-maleimide in DMF containing 2% N-methylmorpholine (NMM) by taking advantage of the free cysteine side chain [65]. 

Figure 3 summarizes the steps involved in cyclic peptide synthesis and these are described in more detail in Subheadings 3.1 and 3.2 for Boc and Fmoc methods, respectively. Carry out all procedures at room temperature and in a well-ventilated fume hood unless otherwise specified. The amounts of reagents are given based on a 0.5 mmol scale synthesis, which typically yields >10 mg of final product after purification. These procedures have been used in our laboratory to synthesize several classes of disulfide-rich cyclic peptides, including SFTI-1 (14 amino acids, one disulfide bond), cyclic Vcl.l (22 amino acid, two disulfide bonds), kalata B1 (29 amino acids, three disulfide bonds), MCoTI-II (34 amino acids, three disulfide bonds), and cyclic chlorotoxin (43 amino acids, four disulfide bonds) (Fig. 4). The procedures are described for manual synthesis but are also applicable to automated SPPS.

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